Intestinal tissue-resident T cell activation depends on metabolite availability.

The metabolic capacity of many cells is tightly regulated and can adapt to changes in metabolic resources according to environmental changes. Tissue-resident memory (TRM) CD8+ T cells are one of the most abundant T cell populations and offer rapid protection against invading pathogens, especially at the epithelia. TRM cells metabolically adapt to their tissue niche, such as the intestinal epithelial barrier. In the small intestine, the types of TRM cells are intraepithelial lymphocytes (IELs), which contain high levels of cytotoxic molecules and express activation markers, suggesting a heightened state of activation. We hypothesize that the tissue environment may determine IEL activity. We show that IEL activation, in line with its semiactive status, is metabolically faster than circulating CD8+ T cells. IEL glycolysis and oxidative phosphorylation (OXPHOS) are interdependently regulated and are dependent on rapid access to metabolites from the environment. IELs are restrained by local availability of metabolites, but, especially, glucose levels determine their activity. Importantly, this enables functional control of intestinal TRM cells by metabolic means within the fragile environment of the intestinal epithelial barrier.

Improving human mesenchymal stem cell-derived hepatic cell energy metabolism by manipulating glucose homeostasis and glucocorticoid signaling.

Introduction: The development of reliable hepatic in vitro models may provide insights into disease mechanisms, linking hepatocyte dysmetabolism and related pathologies. However, several of the existing models depend on using high concentrations of hepatocyte differentiation-promoting compounds, namely glucose, insulin, and dexamethasone, which is among the reasons that have hampered their use for modeling metabolism-related diseases. This work focused on modulating glucose homeostasis and glucocorticoid concentration to improve the suitability of a mesenchymal stem-cell (MSC)-derived hepatocyte-like cell (HLC) human model for studying hepatic insulin action and disease modeling. Methods: We have investigated the role of insulin, glucose and dexamethasone on mitochondrial function, insulin signaling and carbohydrate metabolism, namely AKT phosphorylation, glycogen storage ability, glycolysis and gluconeogenesis, as well as fatty acid oxidation and bile acid metabolism gene expression in HLCs. In addition, we evaluated cell morphological features, albumin and urea production, the presence of hepatic-specific markers, biotransformation ability and mitochondrial function. Results: Using glucose, insulin and dexamethasone levels close to physiological concentrations improved insulin responsiveness in HLCs, as demonstrated by AKT phosphorylation, upregulation of glycolysis and downregulation of Irs2 and gluconeogenesis and fatty acid oxidation pathways. Ammonia detoxification, EROD and UGT activities and sensitivity to paracetamol cytotoxicity were also enhanced under more physiologically relevant conditions. Conclusion: HLCs kept under reduced concentrations of glucose, insulin and dexamethasone presented an improved hepatic phenotype and insulin sensitivity demonstrating superior potential as an in vitro platform for modeling energy metabolism-related disorders, namely for the investigation of the insulin signaling pathway.

Mitochondrial Biogenesis in Neurons: How and Where.

Neurons rely mostly on mitochondria for the production of ATP and Ca2+ homeostasis. As sub-compartmentalized cells, they have different pools of mitochondria in each compartment that are maintained by a constant mitochondrial turnover. It is assumed that most mitochondria are generated in the cell body and then travel to the synapse to exert their functions. Once damaged, mitochondria have to travel back to the cell body for degradation. However, in long cells, like motor neurons, this constant travel back and forth is not an energetically favourable process, thus mitochondrial biogenesis must also occur at the periphery. Ca2+ and ATP levels are the main triggers for mitochondrial biogenesis in the cell body, in a mechanism dependent on the Peroxisome-proliferator-activated γ co-activator-1α-nuclear respiration factors 1 and 2-mitochondrial transcription factor A (PGC-1α-NRF-1/2-TFAM) pathway. However, even though of extreme importance, very little is known about the mechanisms promoting mitochondrial biogenesis away from the cell body. In this review, we bring forward the evoked mechanisms that are at play for mitochondrial biogenesis in the cell body and periphery. Moreover, we postulate that mitochondrial biogenesis may vary locally within the same neuron, and we build upon the hypotheses that, in the periphery, local protein synthesis is responsible for giving all the machinery required for mitochondria to replicate themselves.

The PINK1-Mediated Crosstalk between Neural Cells and the Underlying Link to Parkinson's Disease.

Mitochondrial dysfunction has a fundamental role in the development of idiopathic and familiar forms of Parkinson's disease (PD). The nuclear-encoded mitochondrial kinase PINK1, linked to familial PD, is responsible for diverse mechanisms of mitochondrial quality control, ATP production, mitochondrial-mediated apoptosis and neuroinflammation. The main pathological hallmark of PD is the loss of dopaminergic neurons. However, novel discoveries have brought forward the concept that a disruption in overall brain homeostasis may be the underlying cause of this neurodegeneration disease. To sustain this, astrocytes and microglia cells lacking PINK1 have revealed increased neuroinflammation and deficits in physiological roles, such as decreased wound healing capacity and ATP production, which clearly indicate involvement of these cells in the physiopathology of PD. PINK1 executes vital functions within mitochondrial regulation that have a detrimental impact on the development and progression of PD. Hence, in this review, we aim to broaden the horizon of PINK1-mediated phenotypes occurring in neurons, astrocytes and microglia and, ultimately, highlight the importance of the crosstalk between these neural cells that is crucial for brain homeostasis.

Transcription- and phosphorylation-dependent control of a functional interplay between XBP1s and PINK1 governs mitophagy and potentially impacts Parkinson disease pathophysiology.

Parkinson disease (PD)-affected brains show consistent endoplasmic reticulum (ER) stress and mitophagic dysfunctions. The mechanisms underlying these perturbations and how they are directly linked remain a matter of questions. XBP1 is a transcription factor activated upon ER stress after unconventional splicing by the nuclease ERN1/IREα thereby yielding XBP1s, whereas PINK1 is a kinase considered as the sensor of mitochondrial physiology and a master gatekeeper of mitophagy process. We showed that XBP1s transactivates PINK1 in human cells, primary cultured neurons and mice brain, and triggered a pro-mitophagic phenotype that was fully dependent of endogenous PINK1. We also unraveled a PINK1-dependent phosphorylation of XBP1s that conditioned its nuclear localization and thereby, governed its transcriptional activity. PINK1-induced XBP1s phosphorylation occurred at residues reminiscent of, and correlated to, those phosphorylated in substantia nigra of sporadic PD-affected brains. Overall, our study delineated a functional loop between XBP1s and PINK1 governing mitophagy that was disrupted in PD condition.Abbreviations: 6OHDA: 6-hydroxydopamine; baf: bafilomycin A1; BECN1: beclin 1; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CASP3: caspase 3; CCCP: carbonyl cyanide chlorophenylhydrazone; COX8A: cytochrome c oxidase subunit 8A; DDIT3/CHOP: DNA damage inducible transcript 3; EGFP: enhanced green fluorescent protein; ER: endoplasmic reticulum; ERN1/IRE1α: endoplasmic reticulum to nucleus signaling 1; FACS: fluorescence-activated cell sorting; HSPD1/HSP60: heat shock protein family D (Hsp60) member 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MFN2: mitofusin 2; OPTN: optineurin; PD: Parkinson disease; PINK1: PTEN-induced kinase 1; PCR: polymerase chain reaction:; PRKN: parkin RBR E3 ubiquitin protein ligase; XBP1s [p-S61A]: XBP1s phosphorylated at serine 61; XBP1s [p-T48A]: XBP1s phosphorylated at threonine 48; shRNA: short hairpin RNA, SQSTM1/p62: sequestosome 1; TIMM23: translocase of inner mitochondrial membrane 23; TM: tunicamycin; TMRM: tetramethyl rhodamine methylester; TOMM20: translocase of outer mitochondrial membrane 20; Toy: toyocamycin; TP: thapsigargin; UB: ubiquitin; UB (S65): ubiquitin phosphorylated at serine 65; UPR: unfolded protein response, XBP1: X-box binding protein 1; XBP1s: spliced X-box binding protein 1.

PINK1: A Bridge between Mitochondria and Parkinson's Disease.

Mitochondria are known as highly dynamic organelles essential for energy production. Intriguingly, in the recent years, mitochondria have revealed the ability to maintain cell homeostasis and ultimately regulate cell fate. This regulation is achieved by evoking mitochondrial quality control pathways that are capable of sensing the overall status of the cellular environment. In a first instance, actions to maintain a robust pool of mitochondria take place; however, if unsuccessful, measures that lead to overall cell death occur. One of the central key players of these mitochondrial quality control pathways is PINK1 (PTEN-induce putative kinase), a mitochondrial targeted kinase. PINK1 is known to interact with several substrates to regulate mitochondrial functions, and not only is responsible for triggering mitochondrial clearance via mitophagy, but also participates in maintenance of mitochondrial functions and homeostasis, under healthy conditions. Moreover, PINK1 has been associated with the familial form of Parkinson's disease (PD). Growing evidence has strongly linked mitochondrial homeostasis to the central nervous system (CNS), a system that is replenished with high energy demanding long-lasting neuronal cells. Moreover, sporadic cases of PD have also revealed mitochondrial impairments. Thus, one could speculate that mitochondrial homeostasis is the common denominator in these two forms of the disease, and PINK1 may play a central role in maintaining mitochondrial homeostasis. In this review, we will discuss the role of PINK1 in the mitochondrial physiology and scrutinize its role in the cascade of PD pathology.

Orchestrating mitochondria in neurons: Cytoskeleton as the conductor.

Mitochondria are crucial to support synaptic activity, particularly through ATP production and Ca2+ homeostasis. This implies that mitochondria need to be well distributed throughout the different neuronal sub-compartments. To achieve this, a tight and precise regulation of several neuronal cytoskeleton players is necessary to transport and dock mitochondria. As post-mitotic cells, neurons are highly dependent on mitochondrial quality control mechanisms and several cytoskeleton proteins have been implicated in mitophagy. Therefore, all of these processes are orchestrated by the crosstalk between mitochondria and the neuronal cytoskeleton to form a coordinated and tuned symphony.

A Systematic Pan-Cancer Analysis of Genetic Heterogeneity Reveals Associations with Epigenetic Modifiers.

Intratumor genetic heterogeneity (ITH) is the main obstacle to effective cancer treatment and a major mechanism of drug resistance. It results from the continuous evolution of different clones of a tumor over time. However, the molecular features underlying the emergence of genetically-distinct subclonal cell populations remain elusive. Here, we conducted an exhaustive characterization of ITH across 2807 tumor samples from 16 cancer types. Integration of ITH scores and somatic variants detected in each tumor sample revealed that mutations in epigenetic modifier genes are associated with higher ITH levels. In particular, genes that regulate genome-wide histone and DNA methylation emerged as being determinant of high ITH. Indeed, the knockout of histone methyltransferase SETD2 or DNA methyltransferase DNMT3A using the CRISPR/Cas9 system on cancer cells led to significant expansion of genetically-distinct clones and culminated in highly heterogeneous cell populations. The ITH scores observed in knockout cells recapitulated the heterogeneity levels observed in patient tumor samples and correlated with a better mitochondrial bioenergetic performance under stress conditions. Our work provides new insights into tumor development, and discloses new drivers of ITH, which may be useful as either predictive biomarkers or therapeutic targets to improve cancer treatment.

PARL deficiency in mouse causes Complex III defects, coenzyme Q depletion, and Leigh-like syndrome.

The mitochondrial intramembrane rhomboid protease PARL has been implicated in diverse functions in vitro, but its physiological role in vivo remains unclear. Here we show that Parl ablation in mouse causes a necrotizing encephalomyelopathy similar to Leigh syndrome, a mitochondrial disease characterized by disrupted energy production. Mice with conditional PARL deficiency in the nervous system, but not in muscle, develop a similar phenotype as germline Parl KOs, demonstrating the vital role of PARL in neurological homeostasis. Genetic modification of two major PARL substrates, PINK1 and PGAM5, do not modify this severe neurological phenotype. Parl-/- brain mitochondria are affected by progressive ultrastructural changes and by defects in Complex III (CIII) activity, coenzyme Q (CoQ) biosynthesis, and mitochondrial calcium metabolism. PARL is necessary for the stable expression of TTC19, which is required for CIII activity, and of COQ4, which is essential in CoQ biosynthesis. Thus, PARL plays a previously overlooked constitutive role in the maintenance of the respiratory chain in the nervous system, and its deficiency causes progressive mitochondrial dysfunction and structural abnormalities leading to neuronal necrosis and Leigh-like syndrome.